Volume 98, Issue 5, Pages 597-608 (September 1999) SOCS1 Is a Critical Inhibitor of Interferon γ Signaling and Prevents the Potentially Fatal Neonatal Actions of this Cytokine  Warren S Alexander, Robyn Starr, Jennifer E Fenner, Clare L Scott, Emanuela Handman, Naomi S Sprigg, Jason E Corbin, Ann L Cornish, Rima Darwiche, Catherine M Owczarek, Thomas W.H Kay, Nicos A Nicola, Paul J Hertzog, Donald Metcalf, Douglas J Hilton  Cell  Volume 98, Issue 5, Pages 597-608 (September 1999) DOI: 10.1016/S0092-8674(00)80047-1

Figure 1 Experimentally Unmanipulated SOCS1−/− Mice Show Evidence of an Ongoing Response to IFNγ (A) EMSA of extracts of livers from SOCS1+/− mice injected with 2 μg of IFNγ or untreated SOCS1+/+ and SOCS1−/− mice. Prior to DNA binding, samples were treated, as indicated, with antibodies to either STAT1 or STAT3. (B) Northern blot analyses showing mRNA from brain (b), kidney (k), liver (li), lung (lu), spleen (s), and thymus (t) from SOCS1+/+ and SOCS1−/− mice hybridized with an iNOS probe (upper panel), IRF1 probe (central panel), or GAPDH probe (lower panel). (C) Immunohistochemistry showing class I MHC expression in the liver. Liver sections were stained with anti-class I MHC antibody or isotype control antibody and a peroxidase-labeled secondary antibody. Sections were counterstained with Giemsa. (D) FACS analysis of class I MHC and Mac1 expression on bone marrow cells from SOCS1+/+ and SOCS1−/− mice. Cells were stained with a biotinylated anti-class I MHC antibody and streptavidin-phycoerythrin followed by an FITC-conjugated anti-Mac1 antibody and were analyzed by flow cytometry. Data shown are representative of at least three independent experiments. Cell 1999 98, 597-608DOI: (10.1016/S0092-8674(00)80047-1)

Figure 2 Development of Disease in SOCS1−/− Mice Is Dependent on IFNγ and Occurs in a Germ-Free Environment (A) Rapid onset of morbidity of untreated SOCS1−/− but not SOCS1+/+ or SOCS1+/− mice. (B) Twice weekly injection of neonatal SOCS1−/− mice with anti-IFNγ but not anti-IL-6 or control antibody prevents morbidity. (C) Rapid onset of morbidity in SOCS1−/− but not SOCS1+/+ or SOCS1+/− mice born and reared in a germ-free environment. (D) Prevention of morbidity in SOCS1−/−IFNγ−/− mice and amelioration of morbidity in SOCS1−/−IFNγ+/− mice in comparison with SOCS1−/−IFNγ+/+ mice. Seventeen SOCS1−/−IFNγ−/− mice have been examined to date, with twelve animals being sacrificed for detailed histological and hematological analysis at 3 weeks of age and the remaining five being left for long-term observation. Cell 1999 98, 597-608DOI: (10.1016/S0092-8674(00)80047-1)

Figure 3 Prevention of Tissue Pathology in SOCS1−/− Mice by Treatment with Anti-IFNγ Antibody Neonatal SOCS1−/− mice were treated twice weekly with either anti-IFNγ antibody or control antibody. Anti-IFNγ antibody–treated mice were healthy at 21 days of age and were sacrificed. Control antibody–treated mice developed disease between 9 and 16 days of age and were sacrificed when moribund. The normal histological appearance of organs from anti-IFNγ antibody-treated SOCS1−/− mice (right-hand panels) is contrasted with the atrophy of the cortex of the thymus, the fatty degeneration, hematopoietic infiltration and necrosis of the liver, macrophage accumulation around the bronchi and in the alveolar walls in the lung, and macrophage infiltration and acinar tissue destruction of the pancreas observed in SOCS1−/− mice treated with control antibody (left-hand panels). Cell 1999 98, 597-608DOI: (10.1016/S0092-8674(00)80047-1)

Figure 4 Prevention of B Lymphopenia in SOCS1−/− Mice by Treatment with Anti-IFNγ Antibody or Generation of SOCS1−/−IFNγ−/− Mice Spleen cells were collected from (A) wild-type mice, (B) SOCS1−/− mice treated with a control antibody, (C) SOCS1−/− mice treated with anti-IFNγ antibody, and (D) SOCS1−/−IFNγ−/− mice. Cells were stained with anti-B220 antibody and a mixture of anti-IgM and anti-IgD antibodies and analyzed by FACS. Cell 1999 98, 597-608DOI: (10.1016/S0092-8674(00)80047-1)

Figure 5 Absence of SOCS1 Renders Cells Hyperresponsive to IFNγ and Mice Hyperresistant to Semliki Forest Virus Infection (A) Macrophages from the bone marrow of SOCS1+/+ (filled symbols) and SOCS1−/− mice (open symbols) were stimulated with 100 ng/ml LPS and the indicated concentration of IFNγ and infected with L. major. After 2 hr (circles) and 48 hr (squares), the percentage of macrophages containing parasites was determined. (B) Mice generated by crossing SOCS1+/− mice were infected with Semliki forest virus at 5 days of age and monitored. Cell 1999 98, 597-608DOI: (10.1016/S0092-8674(00)80047-1)